Cancer is one of the leading causes of death. It is a class of diseases which is caused by malignant transformation of healthy cells, caused by genetic alterations, like chromosomal translocations, mutations in tumor suppressor genes, transcription factors or growth-factor receptors, leading to the immortalization of the cells. If the immortalization is combined with excessive proliferation the immortalized cells generate tumors with or without metastasis (in case of solid tumors) or leukemias and lymphomas (cancers of the blood). Defective apoptosis, or programmed cell death, can further contribute to malignant transformation of cells leading to cancer.
A family of membrane associated receptor tyrosine kinases, consisting of the receptor tyrosine kinase orphan receptors-1 and -2 (ROR1 and ROR2) have been described as being specifically associated with particular cancers (Rebagay et al. (2012) Front Oncol. 2(34)), while being largely absent in expression on healthy tissue with few exceptions (Balakrishnan et al. (2016) Clin Cancer Res. doi: 10.1158/1078-0432). Whether or not ROR expression is functionally associated with tumorigenesis remains unclear. However, due to the very tumor-selective expression of the ROR family members, they represent relevant targets for targeted cancer therapies. Receptor tyrosine kinase orphan receptors-1 (ROR1) is of particular interest as a cancer target, due to its nearly 100% association with chronic lymphocytic leukemia (CLL) (Cui et al. (2016) Blood 128(25), p. 2931) and the observation that is also expressed in certain solid tumors, like that of lung and breast (Balakrishnan et al. (2016) Clin Cancer Res. doi: 10.1158/1078-0432). Members of the ROR family are type-I transmembrane proteins containing three distinct extracellular domains, an Ig, a Kringle and a Frizzled domain, followed a transmembrane spanning region, and an intracellular portion. Within the intracellular portion, ROR1 possesses a tyrosine kinase domain, two serine/threonine-rich domains and a proline-rich domain. RORs have been studied in the context of embryonic patterning and neurogenesis through a variety of homologs. These physiologic functions are dichotomous based on the requirement of the kinase domain. A growing literature has established ROR1 as a marker for cancer, such as in chronic lymphocytic leukemia (CLL) for which ROR1 expression is nearly 100% correlated, some acute lymphoblastic leukemias (ALL), mantle cell lymphomas, and some other blood malignancies. In addition, ROR1 is critically involved in progression of a number of solid tumors, such as in neuroblastoma, sarcoma, renal cell carcinoma, breast cancer, lung cancer, colon cancer, head and neck cancer, melanoma, and other cancers. ROR1 has been shown to inhibit apoptosis, potentiate EGFR signaling, induce epithelial-mesenchymal transition (EMT), and contribute to caveolae formation. Importantly, ROR1 is mainly detectable in embryonic tissue and generally absent in adult tissue, making the protein an ideal drug target for cancer therapy. As such, ROR1 has previously been recognized as a target for the development of ROR1 specific antibodies. However, due to the high homology of ROR1 between different mammalian species, which is 100% conserved on the amino acid level between humans and cynomolgus monkeys, 96.7% homologous between human and mouse, and 96.3% homologous between human and rabbit, it has been difficult to raise high affinity antibodies against this target by standard technologies, like animal immunizations.
A few murine and rabbit antibodies have been discussed in the literature. For example, WO 2007/051077 discussed monoclonal antibodies, including humanized antibodies, directed against native ROR1 found on lymphomas including CLL, small lymphocytic lymphoma, marginal B-cell lymphoma and Burkett's lymphoma. Methods for inhibiting growth of a tumor cell using agents, which may be ROR1-binding antibodies that inhibit ROR1 kinase activity, are the subject of WO 2007/146957. WO 2011/054007 discussed a method of treatment or prophylaxis of cancer in which the extracellular domain of ROR1 is expressed by administration of specific ROR1-targeting antibodies.
Additionally, WO 2010/124188 discussed anti-human ROR1 antibodies, and in particular to monoclonal murine antibody referred to under the name 2A2, while WO 2012/075158 refers to monoclonal rabbit antibodies named R11 and R12. Particular ROR1-targeting antibodies are also mentioned in WO 2016/094873. Both WO 2011/079902 and WO 2012/076066 discussed biological inhibitors of ROR1 capable of inducing cell death that bind to selected extracellular ROR1 domain sequences. WO 2014/031174 refers to anti-ROR1 antibodies having the same binding specificity as an antibody named 99961. Binding epitopes of anti-ROR1 antibodies are further referred to in WO 2016/187220. WO 2011/159847 discussed particular scFv antibody fragment conjugates that bind ROR1. WO 2014/167022, WO 2016/055592 and WO 2016/055593 discussed bispecific ROR1-targeting antibodies and their uses, while WO 2015/184203 discussed tri-specific binding molecules. Especially newer documents disclosing humanized anti-ROR1 monoclonal antibodies are based on the originally disclosed mouse or rabbit antibodies, like 2A2, R11, R12 or D10.
Due to the low number of available ROR1 specific monoclonal antibodies, there is a need in the art for better anti-ROR1 antibodies that have higher affinity or other functional properties not possessed by the known antibody clones. There is also a need for additional diagnostic tools for detecting ROR1 expressions in ROR1-related disease conditions by, e.g., Western blotting and/or immunohistochemistry (IHC). The instant invention is directed to addressing these and other needs.